Supplementary Figure 1: Cross-database comparison and experimental setup for preparation and characterization of m6A synaptic RNA in synaptosomes.
a. Summary of cross-database comparison of m6A mRNA and localized mRNA lists. b. Synaptosome purification from healthy adult mouse forebrains using percoll-sucrose discontinuous gradients. F3 and F4 fractions were pooled as synaptosomal fraction (SYN). Homogenate lysate without fractionation was used as control for comparison (HOM); c. Relative (de)enrichment of a panel of synaptic and somatic markers probed using western blot (n=3 independent experiments); d. Full scan of blots in c (n=3 independent experiments); e. Separation of RNA populations was confirmed by qRT-PCR examination of nuclear lncRNA Malat1, somatic RNA beta-3 tubulin and known synaptically localized transcript CaMKIIα (mean value ±s.e.m., n=3 independent experiments). f. Bioanalyzer analysis of RNA integrity in prepared fractions. HOM, homogenate; Cyto, cytoplasm, F3 and F4 (SYN), synaptosomes (n=3 independent experiments). g. A biological replicate of m6A dot-blot presented in Fig. 1b shows highly similar results (n=2 independent experiments).
